Comparison of phenol-chloroform and a commercial deoxyribonucleic acid extraction kit for identification of bloodmeal sources from triatomines (Hemiptera: Reduviidae).

INTRODUCTION: Knowledge of triatomine bloodmeal sources is essential for understanding vector-host interactions in Trypanosoma cruzi transmission cycles. Expensive commercial deoxyribonucleic acid (DNA) extraction kits are widely used for bloodmeal identification. This study assessed the performance of an inexpensive phenol-chloroform DNA extraction protocol for identification of triatomine bloodmeal sources, comparing it with a commercially available kit. METHODS: Both methods were used to obtain DNA from the intestinal contents of Triatoma brasiliensis blood-fed on either Columba sp., Mus musculus, or Gallus gallus. Subsequently, the mitochondrial 12S ribosomal ribonucleic acid (rRNA) gene was amplified by polymerase chain reaction, sequenced, and compared with GenBank data. RESULTS: Twelve (80%) samples extracted with the commercial kit and four (26.7%) with phenol-chloroform were pure (according to the A260/A280 ratio). Samples extracted with phenol-chloroform, except for Columba sp. samples, had higher DNA concentration than those extracted with the commercial kit. Samples extracted using phenol-chloroform and blood-fed on G. gallus had significantly higher DNA concentration than those blood-fed on Columba sp. (p-value <0.001) and M. musculus (p-value <0.001). The 215-base-pair 12S rRNA fragment was amplified from all samples and produced reliable sequences, enabling the identification of the bloodmeal source, most of which showed identity and coverage above 95%. The phenol-chloroform method was much less expensive than the commercial kit but took considerably more time to perform. CONCLUSIONS: Our data showed that both DNA extraction methods produced reliable sequences enabling identification of triatomine bloodmeal sources but differed greatly in cost and time required.

Epidemiological profile of acute Chagas disease in individuals infected by oral transmission in northern Brazil.

INTRODUCTION: Oral infection by Trypanosoma cruzi is currently the most important route of transmission of acute Chagas disease (ACD) in the North region of Brazil, and the reported outbreaks are usually related to ingestion of contaminated food, especially unprocessed açaí pulp. METHODS: A retrospective cohort study was performed to analyze the epidemiological profile of individuals with suspected cases of ACD in the municipality of Breves, located in the state of Pará, Brazil. Therefore, notifications of suspected cases of ACD were collected from the Municipal Health Department of Breves from January 2007 to December 2017. RESULTS: A total of 265 individuals were registered, and the majority were male (54.7%; 145/265). Age ranged from nine months to 79 years, with a greater number of notifications for individuals aged between 1 and 39 years (71.3%; 189/265). Most of them had a low level of education (74.3%, 197/265), were living in rural and urban areas (58.9%; 156/265 and 37.7%; 100/265, respectively). Infection occurred mainly in the domestic environment (96.2%; 255/265) through oral transmission (98.1%; 260/265). There were a greater number of notifications in November, December and January. CONCLUSIONS: These data showed that oral transmission of T. cruzi has become increasingly high in the study region, and health education programs need to be implemented as strategies to ensure good manufacturing practices of unprocessed food.

Epidemiological profile of acute Chagas disease in individuals infected by oral transmission in northern Brazil

Abstract INTRODUCTION: Oral infection by Trypanosoma cruzi is currently the most important route of transmission of acute Chagas disease (ACD) in the North region of Brazil, and the reported outbreaks are usually related to ingestion of contaminated food, especially unprocessed açaí pulp. METHODS A retrospective cohort study was performed to analyze the epidemiological profile of individuals with suspected cases of ACD in the municipality of Breves, located in the state of Pará, Brazil. Therefore, notifications of suspected cases of ACD were collected from the Municipal Health Department of Breves from January 2007 to December 2017. RESULTS A total of 265 individuals were registered, and the majority were male (54.7%; 145/265). Age ranged from nine months to 79 years, with a greater number of notifications for individuals aged between 1 and 39 years (71.3%; 189/265). Most of them had a low level of education (74.3%, 197/265), were living in rural and urban areas (58.9%; 156/265 and 37.7%; 100/265, respectively). Infection occurred mainly in the domestic environment (96.2%; 255/265) through oral transmission (98.1%; 260/265). There were a greater number of notifications in November, December and January. CONCLUSIONS These data showed that oral transmission of T. cruzi has become increasingly high in the study region, and health education programs need to be implemented as strategies to ensure good manufacturing practices of unprocessed food.

Comparison of phenol-chloroform and a commercial deoxyribonucleic acid extraction kit for identification of bloodmeal sources from triatomines (Hemiptera: Reduviidae)

Abstract INTRODUCTION: Knowledge of triatomine bloodmeal sources is essential for understanding vector-host interactions in Trypanosoma cruzi transmission cycles. Expensive commercial deoxyribonucleic acid (DNA) extraction kits are widely used for bloodmeal identification. This study assessed the performance of an inexpensive phenol-chloroform DNA extraction protocol for identification of triatomine bloodmeal sources, comparing it with a commercially available kit. METHODS: Both methods were used to obtain DNA from the intestinal contents of Triatoma brasiliensis blood-fed on either Columba sp., Mus musculus, or Gallus gallus. Subsequently, the mitochondrial 12S ribosomal ribonucleic acid (rRNA) gene was amplified by polymerase chain reaction, sequenced, and compared with GenBank data. RESULTS: Twelve (80%) samples extracted with the commercial kit and four (26.7%) with phenol-chloroform were pure (according to the A260/A280 ratio). Samples extracted with phenol-chloroform, except for Columba sp. samples, had higher DNA concentration than those extracted with the commercial kit. Samples extracted using phenol-chloroform and blood-fed on G. gallus had significantly higher DNA concentration than those blood-fed on Columba sp. (p-value <0.001) and M. musculus (p-value <0.001). The 215-base-pair 12S rRNA fragment was amplified from all samples and produced reliable sequences, enabling the identification of the bloodmeal source, most of which showed identity and coverage above 95%. The phenol-chloroform method was much less expensive than the commercial kit but took considerably more time to perform. CONCLUSIONS: Our data showed that both DNA extraction methods produced reliable sequences enabling identification of triatomine bloodmeal sources but differed greatly in cost and time required.